of Work: Recent instrumental developments in mass spectrometry, such as matrix-assisted laser desorption and electrospray ionization, have enabled mass spectrometrists to investigate biological molecules with significantly higher Mr than previously possible. The combination of these techniques with chemical processing of large biopolymers such as proteins now enables the use of mass spectrometry to play a significant role in the realm of structural biology. In this respect, structural biology not only refers to the determination of the primary sequence and sites of post-translational modifications, but also to probing the tertiary structure of molecules and complexes. We are currently working on several projects, two of which will be described here. The first project is to develop the capability of probing non-covalent interactions between proteins and between proteins and DNA using mass spectrometry. We have successfully investigated the trypsin:BPTI complex using both ESI and MALDI. We have found that, using different instruments and different solution conditions, identical differences in mean charge and in charge envelopes between trypsin and complexed trypsin were observed. This implies that the differences observed are intrinsic to the structural differences between the two states. We are currently investigating differences in surface accessibility and charged residues to understand the data. Another project in this area is the investigation of non-covalent complexes between proteins and DNA. The model system we are working with is a leucine-zipper mimic with oligonucleotide dimers (up to 39-mers). If successful, this will permit investigation of more involved complexes of DNA and proteins relevant to DNA enzymology. We hope, eventually, to include DNA:protein footprinting experiments into this project.A second project is to determine the structure of the human p53 tumor suppressor protein post-translationally modified in response to radiation induced DNA damage (in collaboration with Merrick/Selkirk, LMC). Cellular response to DNA damage is partially mediated through the cell cycle arrest.
Showing the most recent 10 out of 25 publications
